高效液相色谱法测定婴幼儿配方奶粉中1,3-二油酸-2-棕榈酸甘油三酯

摘要/Abstract

摘要： 建立了婴幼儿配方乳粉中1,3-二油酸-2-棕榈酸甘油三酯(1,3-Dioleyl-2-palmitoyl-glycerol，OPO)的检测方法。样品经氨水水解，用有机溶剂提取脂肪，氨基固相萃取柱净化，使用银离子色谱柱分离，以0.55%乙腈-正己烷为流动相洗脱，用高效液相色谱-蒸发光散射检测器(High performance liquid chromatography-evaporative light scattering detector，HPLC-ELSD)检测。该方法实现了OPO和1,2-二油酸-3-棕榈酸甘油三酯(1,2-Dioleyl-3-palmitoyl-glycerol，OOP)的基线分离，可对OPO进行准确定性定量分析；方法在25~500 μg/mL浓度范围内线性关系良好，相关系数R2=0.9996，检出限(LOD)和定量限(LOQ)为0.30、0.90 g/kg，加标浓度为1~96 g/kg的范围内，OPO的平均回收率在97.1%~104.2%之间，RSD为1.1%~2.9%，精密度和正确度等方法学指标均符合要求；还通过了实验室间的协同性验证。调查分析了我国市售39份OPO强化婴幼儿配方奶粉，OPO含量仅为标签标示量的28.4%~59.7%，这主要是因为检测方法不一致所造成的。

关键词: 高效液相色谱-蒸发光散射检测器, 1,3-油酸-2-棕榈酸甘油三酯, 婴幼儿配方奶粉

Abstract: A method for detecting 1,3-dioleyl-2-palmitoyl-glycerol (OPO) in infant formula has been established. The sample is hydrolyzed with ammonia, and the fat is extracted with an organic solvent. Amino solid-phase extraction column is used for purification, and silver ion chromatography column is used for separation. Elution is performed using a mobile phase of 0.55% acetonitrile-hexane, and detection is carried out using high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). This method achieves baseline separation of OPO and 1,2-dioleyl-3-palmitoyl-glycerol (OOP), allowing for accurate quantification of OPO. The method shows good linearity in the concentration range of 25-500 μg/mL, with a correlation coefficient R2= 0.9996. The limit of detection (LOD) and quantification (LOQ) are 0.30 and 0.90 g/kg, respectively. Within the range of spiked concentrations from 1 to 96 g/kg, the average recovery rate of OPO ranges from 97.1% to 104.2%, with relative standard deviations (RSD) between 1.1% and 2.9%. The method meets the requirements for precision and accuracy, and has also passed inter-laboratory collaborative validation. An investigation analyzed 39 commercially available OPO-fortified infant formula milk powders in China, revealing that the actual OPO content ranged from28.4% to 59.7% of the labeled amount. This discrepancy is primarily due to inconsistent detection methods.

Key words: High performance liquid chromatography-evaporative light scattering detector, 1, 3-Oleic acid-2-palmitoyl triglyceride, Infant formula

中图分类号:

TS252.7

戴嵘德刘峰傅武胜黄思婷杨贵芝刘文菁. 高效液相色谱法测定婴幼儿配方奶粉中1,3-二油酸-2-棕榈酸甘油三酯[J]. 食品科学.

rongde DAI FU Wu－Sheng Fu. Determination of 1,3-Dioleic Acid-2-Palmitoyl Triglyceride in Infant Formulae by High Performance Liquid Chromatography[J]. FOOD SCIENCE.

参考文献

[1] KALLIO H, NYLUND M, BOSTROM P, et al.Triacylglycerol regioisomers in human milk resolved with an algorithmic novel electrospray ionization tandem mass spectrometry method[J]. Food Chemistry, 2017, 233:351-360. DOI:10.1016/j.foodchem.2017.04.122.[J].Food Chemistry, 2017, 233(0):351-360
[2] FABRITIUS M, LINDERBORG K M, TARVAINEN M, et al.Direct inlet negative ion chemical ionization tandem mass spectrometric analysis of triacylglycerol regioisomers in human milk and infant formulas[J].[J].Food Chemistry, 2020, 328(0):126991-
[3]ZHANG Xinghe, QI Ce, ZHANG Yiren, et al.Identification and quantification of triacylglycerols in human milk fat using ultra-performance convergence chromatography and quadrupole time-of-flight mass spectrometery with supercritical carbon dioxide as a mobile phase[J].Food Chemistry, 2019, 275(0):712-720
[4]BRONSKY J, CAMPOY C, EMBLETON N, et al.Palm oil and beta-palmitate in infant formula: a position paper by the European Society for Paediatric Gastroenterology,Hepatology,And Nutrition (ESPGHAN) committee on nutrition[J].Journal of Pediatric Gastroenterology and Nutrition, 2019, 68(5):742-760
[5]LITMANOVITZ I, DAVIDSON K, ELIAKIM A, et al.High beta-palmitate formula and bone strength in term infants: a randomized,double-blind,controlled trial[J].Calcified Tissue International, 2013, 92(1):35-41
[6]SHEN Lili, HUANG Weihua, XU Xuebing, et al.Stool saponified fatty acid, behavior, growth, and stool characteristics in infants fed a high-opo formula: a randomized, double-blind clinical trial[J].Frontiers in Pediatrics, 2021, 9(0):-
[7]余章斌，韩树萍，朱春，等.配方奶添加棕榈油对婴儿营养吸收和排便影响的分析[J].中华儿科杂志, 2009, 47(12):904-910
[8]YARON S, SHACHAR D, ABRAMAS L, et al.Effect of high beta-palmitate content in infant formula on the intestinal microbiota of term infants[J].Journal of Pediatric Gastroenterology and Nutrition, 2013, 56(4):376-381
[9]WANG Jing, LIU Lihua, LIU Libo, et al.Absorption of 1,3-dioleyl-2-palmitoylglycerol and intestinal flora profiles changes in mice[J].International Journal of Food Sciences and Nutrition, 2020, 71(3):296-306
[10]郭丹颖.添加sn-2棕榈酸酯的婴儿配方奶粉对婴儿肠道菌群和代谢物的影响[D]. 江南大学, 2022:29-44.
[11]NI Mengmei, WANG Yingyao, WU Rui, et al.Total and sn-2 fatty acid pro fi le in human colostrum and mature breast milk of women living in inland and coastal areas of china.[J].Annals of Nutrition and Metabolism, 2021, 77(1):29-37
[12]董玲，周昕玥，姜毅康，等.母乳中二位棕榈酸酯含量分析及研究现状[J].食品与机械, 2022, 38(9):1-6
[13]WEI Wei, SUN Cong, WANG Xiaosan, et al.Lipase-Catalyzed synthesis of sn-2 palmitate: a review[J].Engineering, 2020, 6(4):406-414
[14]AGOSTONI C, BRESSON J-L, FAIRWEATHER-TAIT S, et al.Scientific opinion on the substantiation of a health claim related to beta‐palmitate and increased calcium absorption pursuant to article 14 of regulation (EC) No 1924/2006[J]. EFSA Journal, 2011, 9(12):2470-
[15]AGOSTONI C, CANANI R B, FAIRWEATHER-TAIT S, et al.Scientific opinion on the substantiation of a health claim related to beta‐palmitate and contribution to softening of stools pursuant to article 14 of regulation (EC) No 1924/2006[J].EFSA Journal, 2014, 12(2):3578-
[16]中华人民共和国卫生部公告(2008年第13号)[EB].中华人民共和国卫生部公报[J].中华人民共和国卫生部公报, 2008, 7(0):6-19
[17]国家卫生健康委员会.食品安全国家标准食品营养强化剂使用标准:GB 14880-2012 [S]. 北京: 中国标准出版社，2012: 10.
[18]国家卫生健康委员会.食品安全国家标准食品营养强化剂 1, 3-二油酸-2-棕榈酸甘油三酯:GB 30604-2015[S]. 北京: 中国标准出版社，2015: 3-5.
[19]陈思宇.银离子色谱柱用于食用油成分分析的研究[J].农业与技术, 2020, 40(6):34-36
[20]韦伟，屠海云，王红青，等.测定, -二油酸--棕榈酸三酰甘油的含量[J].中国粮油学报, 2014, 29(0):105-109
[21]ADLOF R, LIST G.Analysis of triglyceride isomers by silver-ion high-performance liquid chromatography[J].Journal of Chromatography A, 2004, 1046(1-2):109-113
[22]马小宁，田燕，宋小莉，等.气相色谱法测定婴幼儿配方乳粉中, -二油酸-棕榈酸甘油三酯[J].检验检疫学刊, 2013, 23(6):32-38
[23]SASAKI R, UMEZAWA M, TSUKAHARA S, et al.Assignment of milk fat fatty acid propyl esters by gc-fid analysis with the aid of ag-ion solid-phase extraction[J].Journal of Oleo Science, 2015, 64(12):1251-1258
[24]LISA M, VELINSKA H, HOLCAPEK M.Regioisomeric characterization of triacylglycerols using silver-ion hplcms and randomization synthesis of standards[J].Analytical Chemistry (Washington), 2009, 81(10):3903-3910
[25]尚东洋，高杨.气相色谱-质谱联用法测定婴幼儿配方乳粉中1, 3-二油酸-2-棕榈酸甘油三酯含量[J].中国乳业杂志, 2017, 5(0):77-79
[26]HOLCAPEK M, VELINSKA H, LISA M, et al.Orthogonality of silver-ion and non-aqueous reversed-phase HPLCMS in the analysis of complex natural mixtures of triacylglycerols[J].Journal of Separation Science, 2009, 32(21):3672-3680
[27]何榕，山晓琳，董方圆，等.反相超效液相色谱-质谱联用分离分析食用油中的甘油三酯[J].分析化学, 2015, 43(9):1377-1382
[28]宋戈，亢美娟，杨金宝.正相液相色谱法测定婴幼儿配方乳粉中, -二油酸--棕榈酸甘油三酯[J].色谱, 2016, 34(3):351-355
[29]魏婷婷，杨虹，杨天奎.串联银离子色谱柱分析结构甘油三酯同分异构体的研究[J].中国油脂, 2012, 37(07):79-81
[30]GIUFFRIDA F, MARMET C, TAVAZZI I, et al.Quantification of 1,3-olein-2-palmitin (OPO) and palmitic acid in sn-2 position of triacylglycerols in human milk by liquid chromatography coupled with mass spectrometry[J].Molecules, 2019, 24(1):22-
[31]林丽珊，傅武胜，张一可，等.气相色谱-串联质谱法测定婴幼儿配方奶粉中氯丙醇酯和缩水甘油酯[J].食品科学, 2020, 41(22):295-300
[32]国家食品药品监督管理总局, 国家卫生和计划生育委员会.食品安全国家标准食品中脂肪的测定:GB 5009.6-2016[S]. 北京: 中国标准出版社，2016: 5-7.
[33]KURVINEN J P, SJ?VALL O, KALLIO H.Molecular weight distribution and regioisomeric structure of triacylglycerols in some common human milk substitutes[J].Journal of the American Oil Chemists' Society, 2002, 79(1):13-22
[34]VYSSOTSKI M, BLOOR S J, LAGUTIN K, et al.Efficient separation and analysis of triacylglycerols: quantitation of β-palmitate (opo) in oils and infant formulas[J].Journal of Agricultural and Food Chemistry, 2015, 63(26):5985-5992
[35]MOMCHILOVA S M, NIKOLOVA-DAMYANOVA B M.Advances in silver ion chromatography for the analysis of fatty acids and triacylglycerols-2001 to 2011[J].Analytical Sciences, 2012, 28(9):837-844
[36]黄宝勇，欧阳喜辉，潘灿平.色谱法测定农产品中农药残留时的基质效应[J].农药学学报, 2005, 7(4):299-305
[37]中华人民共和国卫生部.食品安全国家标准预包装食品营养标签通则: GB 28050-2011[S]. 北京: 中国标准出版社, 2011: 4.
[38]国家标准化管理委员会, 国家质量监督检验检疫总局.合格评定化学分析方法确认和验证指南: GB/T 27417-2017[S]. 北京: 中国标准出版社, 2017: 4-15.